New fluorimetric assay of horseradish peroxidase using sesamol as substrate and its application to EIA

Horseradish peroxidase (HRP) is generally used as a label enzyme in enzyme immunoassay (EIA). The procedure used for HRP detection in EIA is critical for sensitivity and precision. This paper describes a novel fluorimetric assay for horseradish peroxidase (HRP) using sesamol as substrate. The principle of the assay is as follow: sesamol (3,4-methylenedioxy phenol) is reacted enzymatically in the presence of hydrogen peroxide to produce dimeric sesamol. The dimer is fluorescent and can be detected sensitively at ex. 347 nm, em. 427 nm.The measurable range of HRP was 1.0×10^?18 to 1.0×10^?15 mol/assay, with a detection limit of 1.0×10^?18 mol/assay. The coefficient of variation (CV, n=8) was examined at each point on the standard curve, with a mean CV percentage of 3.8%. This assay system was applied to thyroid stimulating hormone (TSH) EIA using HRP as the label enzyme.     

Efficacy of plasma exchange therapy for Kawasaki disease intractable to intravenous gamma-globulin

Kawasaki disease (KD) causes coronary artery lesions (CALs) in 500 Japanese children each year. Intravenous gamma-globulin (IVGG) decreases the incidence of these lesions from 25% to 8% of the total KD cases. We examined whether plasma exchange is a safe and effective prophylaxis against CALs in children with KD intractable to IVGG therapy. Eighty-nine children with KD at high risk of CALs were selected on the basis of increases in fractional changes in inflammatory markers such as white blood cell count, neutrophil count, and C-reactive protein between the baseline and 1–2 days after IVGG treatment. Of 105 children who received a second course of IVGG therapy because the initial course was ineffective, plasma exchange (PE) was performed in 46 children who had not responded to the second IVGG treatment. The outcome was compared with the results when a third course of IVGG therapy was given to the other 59 children. No complications occurred with the plasma exchange therapy. CALs developed in only 8 of the 46 children (17.3%) who underwent plasma exchange, but they occurred in 24 of the 59 (40.7%) who had received a third course of IVGG therapy (P 0.0012). We concluded that PE was a safe, effective prophylactic measure against CALs in children with KD intractable to IVGG therapy. PE should be performed at an early stage, as soon as fractional increases in inflammatory markers are found after IVGG therapy.     

Pitavastatin, an HMG-CoA reductase inhibitor, exerts eNOS-independent protective actions against angiotensin II induced cardiovascular remodeling and renal insufficiency

Angiotensin II (Ang II) plays a pivotal role in cardiovascular remodeling leading to hypertension, myocardial infarction, and stroke. Pitavastatin, an HMG-CoA reductase inihibitor, is known to have pleiotropic actions against the development of cardiovascular remodeling. The objectives of this study were to clarify the beneficial effects as well as the mechanism of action of pitavastatin against Ang II-induced organ damage. C57BL6/J mice at 10 weeks of age were infused with Ang II for 2 weeks and were simultaneously administered pitavastatin or a vehicle. Pitavastatin treatment improved Ang II-induced left ventricular hypertrophy and diastolic dysfunction and attenuated enhancement of cardiac fibrosis, cardiomyocyte hypertrophy, coronary perivascular fibrosis, and medial thickening. Ang II-induced oxidative stress, cardiac TGFbeta-1 expression, and Smad 2/3 phosphorylation were all attenuated by pitavastatin treatment. Pitavastatin also reduced Ang II-induced cardiac remodeling and diastolic dysfunction in eNOS-/- mice as in wild-type mice. In eNOS-/- mice, the Ang II-induced cardiac oxidative stress and TGF-beta-Smad 2/3 signaling pathway were enhanced, and pitavastatin treatment attenuated the enhanced oxidative stress and the signaling pathway. Moreover, pitavastatin treatment reduced the high mortality rate and improved renal insufficiency in Ang II-treated eNOS-/- mice, with suppression of glomerular oxidative stress and TGF-beta-Smad 2/3 signaling pathway. In conclusion, pitavastatin exerts eNOS-independent protective actions against Ang II-induced cardiovascular remodeling and renal insufficiency through inhibition of the TGF-beta-Smad 2/3 signaling pathway by suppression of oxidative stress.     

Segmental pulmonary vein antrum isolation using the "large-size" lasso catheter in patients with atrial fibrillation.

BACKGROUND: The limited efficacy and complications of segmental ostial pulmonary vein isolation (PVI) for treating atrial fibrillation (AF) have been discussed so, in the present study the feasibility and efficiency of performing segmental pulmonary vein (PV) antrum isolation to treat AF were assessed. METHODS AND RESULTS: A total of 187 patients with drug-refractory AF (paroxysmal 120, persistent 67) underwent segmental PVI guided by circumferential 20-electrode catheters (Lasso). Radiofrequency (RF) current was delivered either at the ostium using a regular Lasso (15-20 mm in diameter, 70 patients: Group 1) or at the antrum using a larger Lasso (25-30 mm in diameter, 117 patients: Group 2). A significantly wider region had to be ablated, with a longer RF application time, to isolate all 4 PVs in Group 2 patients than in Group 1 patients. Although the rate of recurrence of AF after the initial session was equal in both groups, a significantly greater number of patients were free from AF after a mean of 1.4 procedures in Group 2 than in Group 1 (93% vs 76% for paroxysmal AF, 78% vs 48% for persistent AF). CONCLUSIONS: Segmental antral PVI using large-sized Lasso catheters was found to be more effective and safer than ostial PVI for the treatment of AF.     

Role of fibrinolysis in tissue-factor-induced disseminated intravascular coagulation in rats - an effect of tranexamic acid

We clarified the role of fibrinolysis in tissue-factor (TF)-induced rat disseminated intravascular coagulation (DIC) using tranexamic acid (TA). TA suppressed the elevation in D-dimer levels normally observed following TF-induced DIC, and an increase in organ dysfunction was seen. Enhanced fibrinolysis plays an important role in preventing the development of organ failure in TF-induced DIC.     

Genotype,serum level of hepatitis C virus RNA and liver histology as predictors of response to interferon-α 2α therapy in Japanese patients with chronic hepatitis C

Abstract: To determine whether pretreatment HCV-RNA level, hepatitis C virus genotypes, alanine aminotransferase and histology correlate with subsequent response to interferon-α therapy or not, serum HCV-RNA levels and genotype were determined by branched DNA signal amplification assay and genotype-specific polymerase chain reaction in 43 patients with chronic active hepatitis C. Response to recombinant interferon-α 2α (504 million units in total) was defined as complete and sustained CR→SR, n=12), complete response followed by relapse (CR→Rel, n=17), and no response (NR, n=10), excluding dropouts (n=4). Patients who showed CR→SR had a lower HCV-RNA level (0.438 × 10⁶ eq/ml) compared to CR→Rel (2.452 × 10⁶ eq/ml, p=0.008) and NR (4.882 × 10⁶ eq/ml, p=0.009). A higher proportion of patients with CR→SR had type 2a HCV (67%) compared to the CR→Rel (28%) and the NR (0%). There was a trend for type 1b hepatitis C virus infection to have higher serum HCV-RNA levels. There was no correlation between pretreatment HCV-RNA level and alanine aminotransferase. However, no relation between pretreatment HCV-RNA level and liver histology was observed; a high proportion of patients with CAH2a showed CR→SR, compared to those with CAH2b (p=0.001). Moreover, the patients with CAH2b who had low level hepatitis C virus viremia did not show CR→SR. These data indicate that pre-treatment serum HCV-RNA levels, genotype and liver histology are good predictors of subsequent response to interferon-α therapy in Japanese patients with chronic hepatitis C virus infection.     

大鼠降结肠上皮γ-氨基丁酸及谷氨酸脱羧酶的表达特征

目的:检测γ-氨基丁酸(gamma-aminobutyricacid,GABA)和谷氨酸脱羧酶(glutamicaciddecarboxylase,GAD)在大鼠降结肠上皮的表达及分布特征,并探讨GABA与上皮细胞分化增殖的关系.方法:用免疫荧光及激光共聚焦显微扫描技术,检测GABA、GAD65及GAD67在大鼠降结肠上皮中的表达,并以麦芽凝聚素组织化学染色与免疫荧光结合的双重染色显示GABA和GAD65表达细胞的分布特征.同时,用RT-PCR和原位分子杂交方法检测GADmRNA的表达.此外,用3H-胸腺嘧啶放射自显影法显示降结肠上皮的增殖带.结果:RT-PCR显示降结肠黏膜中GAD65及GAD67mRNA均阳性,原位杂交显示阳性杂交信号主要分布在上皮细胞的隐窝和腔面,且GAD65信号较GAD67强.GABA及GAD65免疫反应阳性细胞主要分布在降结肠的腔面和隐窝的上1/3上皮细胞的胞质,而GAD67阳性细胞仅分布腔面,此外,GABA及GAD65阳性染色也见于黏膜固有层.双重染色显示杯状细胞中GABA及GAD65均阴性.3H-胸腺嘧啶标记阳性细胞主要在隐窝的中下段.结论:GABA及GAD65分布在大鼠降结肠上皮的成熟带及功能带,GABA系统可能参与上皮细胞的分化与增殖的调节.